A major gap in chemical modification technologies of proteins are the lack of methods for the site-selective/chemospecific labeling of lysines. Indeed, a general screen for the site-selective labeling of the most accessible lysines in proteins would be advantageous in a number of respects.
First, since most proteins contain numerous lysines it would provide optional, diverse candidates for conjugate development. The candidates would emerge from screens ranked in order of lysine selectivity. For projects which have had a restricted focus on a single target protein site for attachment, this feature is invaluable for diversifying the screening process and bringing it more in line with the “numbers game” of small molecule development. Regioisomers (positional isomers) of purified conjugates would be expected to have different properties (e.g., pharmacokinetic) that could provide insights for clinical development of the optimal conjugate.
Second, labor intensive molecular biology methods are rendered an unnecessary preliminary to introducing payloads onto proteins, as the native or commercially available proteins can be screened without prior modification.
Thirdly, amines can be targeted with diverse acylating entities to provide amide bonds that are among the most chemically stable under physiological conditions of pH and temperature.
Fourthly, affinity elements, and/or labeling entities to the targeted protein could be combined with information on known ligands to site-selectively label of the protein
For these reasons, methods for the site-selective/chemospecific modification and ligation of proteins, and particularly antibodies, would be useful to produce and facilitate the formation of protein conjugates and related crosslinked products such as antibody-drug conjugates. The conjugates and crosslinked products themselves are useful in many respects including molecular diagnostic and therapeutic applications.